The lysine binding sites of human plasminogen. Evidence for a critical tryptophan in the binding site of kringle 4.
Chemical modification of human degraded form of plasminogen with NH2-terminal lysine (Lys-plasminogen) and the elastase fragments kringle 1 + 2 + 3 and kringle 4 with the tryptophan reagent [14C]dimethyl(2-hydroxy-5-nitrobenzyl)sulfonium bromide results in the incorporation of label and the parallel loss of lysine binding ability. In the case of kringle 4, only one-half of the lysine binding sites could be inactivated, but the modified and unmodified forms could be separated by affinity chromatography. The modified form contained 1 mol of 2-hydroxy-5-nitrobenzyl groups/mol of kringle 4 and did not bind to lysine-Sepharose. Lysine analogs such as 6-aminohexanoic acid protected kringle 4 against modification. Peptide-mapping studies on this form showed that essentially all of the label was in two chymotryptic peptides containing a tryptophan corresponding to Trp426 in the plasminogen sequence. Competition experiments with anti-kringle 4 antibodies having an affinity for the lysine binding site showed that the binding of 2-hydroxy-5-nitrobenzyl-kringle 4 to antibodies was about 10 times weaker than for unmodified kringle 4. These results indicate that the integrity of specific tryptophan residue is critical to the binding of lysine and related amino acids to kringle 4of human plasminogen.
D000595:Amino Acid Sequence; D002918:Chymotrypsin; D006801:Humans; D007106:Immune Sera; D007202:Indicators and Reagents; D008239:Lysine; D010196:Pancreatic Elastase; D010446:Peptide Fragments / Q000032:analysis; D010958:Plasminogen / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011863:Radioimmunoassay; D013452:Sulfonium Compounds; D014364:Tryptophan
D007106:Immune Sera; D007202:Indicators and Reagents; D010446:Peptide Fragments; D013452:Sulfonium Compounds; C024586:dimethyl(2-hydroxy-5-nitrobenzyl)sulfonium; D014364:Tryptophan; D010958:Plasminogen; D002918:Chymotrypsin; D010196:Pancreatic Elastase; D008239:Lysine